Steroid structural requirements for interaction of ostreolysin, a lipid-raft binding cytolysin, with lipid monolayers and bilayers |
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Authors: | Katja Rebolj Peter Ma?ek |
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Affiliation: | a Department of Biology, Biotechnical Faculty, University of Ljubljana, Ve?na pot 111, 1000 Ljubljana, Slovenia b Department of Food Science and Technology, Biotechnical Faculty, University of Ljubljana, Jamnikarjeva 101, 1000 Ljubljana, Slovenia |
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Abstract: | Ostreolysin, a cytolytic protein from the edible oyster mushroom (Pleurotus ostreatus), recognizes and binds specifically to membrane domains enriched in cholesterol and sphingomyelin (or saturated phosphatidylcholine). These events, leading to permeabilization of the membrane, suggest that a cholesterol-rich liquid-ordered membrane phase, which is characteristic of lipid rafts, could be its possible binding site. In this work, we present effects of ostreolysin on membranes containing various steroids. Binding and membrane permeabilizing activity of ostreolysin was studied using lipid mono- and bilayers composed of sphingomyelin combined, in a 1/1 molar ratio, with natural and synthetic steroids (cholesterol, ergosterol, β-sitosterol, stigmasterol, lanosterol, 7-dehydrocholesterol, cholesteryl acetate, and 5-cholesten-3-one). Binding to membranes and lytic activity of the protein are both shown to be dependent on the intact sterol 3β-OH group, and are decreased by introducing additional double bonds and methylation of the steroid skeleton or C17-isooctyl chain. The activity of ostreolysin mainly correlates with the ability of the steroids to promote formation of liquid-ordered membrane domains, and is the highest with cholesterol-containing membranes. Furthermore, increasing the cholesterol concentration enhanced ostreolysin binding in a highly cooperative manner, suggesting that the membrane lateral distribution and accessibility of the sterols are crucial for the activity of this new member of cholesterol-dependent cytolysins. |
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Keywords: | 7-DHC, 7-dehydrocholesterol CDC, cholesterol-dependent cytolysin Ch-Ac, cholesteryl acetate Chol, cholesterol Ch-one, 5-cholesten-3-one DOPC, dioleoylphosphatidylcholine DPH, 1,6-diphenyl-1,3,5-hexatriene DPPC, dipalmitoylphosphatidylcholine DRM, detergent-resistant membrane DSC, differential scanning calorimetry EDTA, ethylenediaminetetraacetic acid Ergo, ergosterol Lano, lanosterol ld, liquid disordered domain lo, lipid ordered domain MLV, multilamellar vesicles PC, phosphatidylcholine POPC, palmitoyloleoylphosphatidylcholine RU, resonance unit Sito, β-sitosterol SPR, surface plasmon resonance Stig, stigmasterol SV, sonicated vesicles Oly, ostreolysin SM, sphingomyelin TRIS, (hydroxymethyl)aminomethane VCC, Vibrio cholerae cytolysin |
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